Mutations in homologous recombination (HR) DNA repair genes BRCA1 and BRCA2 predispose to tumorigenesis. HR-deficient cancers are hypersensitive to Poly (ADP ribose)-polymerase (PARP) inhibitors, but can acquire resistance and relapse. The mechanisms underlying PARP inhibitor resistance are only partially understood. This lack of knowledge in part stems from incomplete understanding of how PARP inhibitors induce cytotoxicity in HR-deficient cancers. The aim of this study was to investigate the contribution of mitotic failure to PARP inhibitor-induced cytotoxicity. Using the DNA fiber assay we observed compromised replication fork protection upon BRCA2 depletion, which was aggravated upon PARP inhibition. In accordance with the loss of replication fork stability, we found increased FANCD2 foci in interphase cells upon BRCA2 depletion, and a significant further increase when BRCA2-depleted cells were treated with PARP inhibitor as assessed by immunofluorescence. Importantly, these replication lesions were not resolved prior to mitotic entry, as increased numbers of FANCD2 foci were observed in mitotic cells of BRCA2-depleted cells, which were further increased upon PARP inhibitor treatment. Subsequently we found that in mitosis, BRCA2 or BRCA1 and Brca2 or Brca1 deletion resulted in elevated levels of chromatin-bridges and lagging chromosomes which were greatly enhanced after PARP inhibition. Live-cell microscopy showed that chromatin bridges in anaphase often lead to cytokinesis failure with subsequent multinucleated progeny cells or cell death. Furthermore, PARP inhibitor-induced multinucleated cells failed clonogenic outgrowth and high percentages of multinucleated cells were found in vivo in the remnants of Brca2-/-;p53-/- and Brca1-/-;p53-/- mammary mouse tumors upon PARP inhibitor treatment, suggesting that mitotic progression promotes PARP inhibitor-induced cell death. Indeed, enforced mitotic bypass through EMI1 depletion abrogated the cytotoxic potential of PARP inhibitor sensitivity and indicates that progression through mitosis promotes cell death in response to PARP inhibitors in HR-defective cells. These findings add to our understanding how PARP inhibition induces cytotoxicity. Furthermore, these results provide starting points in tackling tumor resistance to PARP inhibitors, and point at combination therapies which could potentiate the effects of PARP inhibitors for the treatment of HR-deficient tumors.
Citation Format: Pepijn M. Schoonen, Peter Bouwman, Floris Foijer, Madalena Tarsounas, Sohvi Blatter, Jos Jonkers, Sven Rottenberg, Marcel A. Vugt, van. Progression through mitosis promotes PARP inhibitor induced cytotoxicity in homologous recombination deficient cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3453. doi:10.1158/1538-7445.AM2017-3453